Intrathecal agonists for inhibitory G coupled receptors (e.g. mu and alpha2) produce analgesia. Based on receptor distribution and block of afferent release (substance P: SP), we think the analgesia reflects a joint pre- and post-synaptic action. Neurokinin 1 receptor (NK1) internalization and cFos/pCREB expression are robust tools to define in vivo the effect of acute and chronic interventions on small afferent release and post synaptic excitability and to explore several hypothesized ramifications of this action. 1) IT analgesic activity will co-vary with the inhibitory effects on small afferent terminal excitability (NK1 internalization). We will examine in rats the effects/pharmacology of IT agents acting on small afferent terminal receptors (mu, delta, alpha2, NPY, GABAB, and Adenosine A1) to block paw injury evoked NK1 internalization. 2) Spinal receptors located postsynaptic to the primary afferent will control excitatory outflow of spinal neurons driven by afferent input. Using cFos/pCREB expression, paw injury, IT-SP or IT-NMDA will be used to activate spinal neurons and indirectly supraspinal neurons (n. parabrachialis) in the presence of IT-analgesics. 3) Systemic morphine exerts its effect upon spinal function by concurrent spinal and supraspinal actions. We will examine the spinal effects of intracerebral and systemically administered morphine at analgesic doses on spinal NK1 internalization and activation of spinal/supraspinal cFos/pCREB by paw injury, IT SP or IT NMDA. 4) Given tolerance and dependence observed with chronic IT mu and alpha2 agonists and the proposed sensitization of spinal processing mediated though NMDA and PKC, we hypothesize that i) chronic IT mu or alpha2 agonist infusion will enhance injury evoked NK1 internalization and increased IT SP-evoked cFos activation at spinal and supraspinal levels; ii) increased NK1 internalization and cFos after naloxone; and iii) a reversal of mu tolerance with NMDA antagonism or PKC inhibition. In short, these studies seek to illuminate the complex pre- and postsynaptic mechanisms by which modulatory receptors regulate spinal nociceptive outflow.